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Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties

Aptamers are short synthetic DNA or RNA oligonucleotides that adopt secondary and tertiary conformations based on Watson–Crick base-pairing interactions and can be used to target a range of different molecules. Two aptamers, HD1 and HD22, that bind to exosites I and II of the human thrombin molecule...

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Autores principales: Hughes, Quintin W., Le, Bao T., Gilmore, Grace, Baker, Ross I., Veedu, Rakesh N.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6151750/
https://www.ncbi.nlm.nih.gov/pubmed/29048375
http://dx.doi.org/10.3390/molecules22101770
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author Hughes, Quintin W.
Le, Bao T.
Gilmore, Grace
Baker, Ross I.
Veedu, Rakesh N.
author_facet Hughes, Quintin W.
Le, Bao T.
Gilmore, Grace
Baker, Ross I.
Veedu, Rakesh N.
author_sort Hughes, Quintin W.
collection PubMed
description Aptamers are short synthetic DNA or RNA oligonucleotides that adopt secondary and tertiary conformations based on Watson–Crick base-pairing interactions and can be used to target a range of different molecules. Two aptamers, HD1 and HD22, that bind to exosites I and II of the human thrombin molecule, respectively, have been extensively studied due to their anticoagulant potentials. However, a fundamental issue preventing the clinical translation of many aptamers is degradation by nucleases and reduced pharmacokinetic properties requiring higher dosing regimens more often. In this study, we have chemically modified the design of previously described thrombin binding aptamers targeting exosites I, HD1, and exosite II, HD22. The individual aptamers were first modified with an inverted deoxythymidine nucleotide, and then constructed bivalent aptamers by connecting the HD1 and HD22 aptamers either through a triethylene glycol (TEG) linkage or four consecutive deoxythymidines together with an inverted deoxythymidine nucleotide at the 3′-end. The anticoagulation potential, the reversal of coagulation with different antidote sequences, and the nuclease stability of the aptamers were then investigated. The results showed that a bivalent aptamer RNV220 containing an inverted deoxythymidine and a TEG linkage chemistry significantly enhanced the anticoagulation properties in blood plasma and nuclease stability compared to the existing aptamer designs. Furthermore, a bivalent antidote sequence RNV220AD efficiently reversed the anticoagulation effect of RNV220 in blood plasma. Based on our results, we believe that RNV220 could be developed as a potential anticoagulant therapeutic molecule.
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spelling pubmed-61517502018-11-13 Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties Hughes, Quintin W. Le, Bao T. Gilmore, Grace Baker, Ross I. Veedu, Rakesh N. Molecules Article Aptamers are short synthetic DNA or RNA oligonucleotides that adopt secondary and tertiary conformations based on Watson–Crick base-pairing interactions and can be used to target a range of different molecules. Two aptamers, HD1 and HD22, that bind to exosites I and II of the human thrombin molecule, respectively, have been extensively studied due to their anticoagulant potentials. However, a fundamental issue preventing the clinical translation of many aptamers is degradation by nucleases and reduced pharmacokinetic properties requiring higher dosing regimens more often. In this study, we have chemically modified the design of previously described thrombin binding aptamers targeting exosites I, HD1, and exosite II, HD22. The individual aptamers were first modified with an inverted deoxythymidine nucleotide, and then constructed bivalent aptamers by connecting the HD1 and HD22 aptamers either through a triethylene glycol (TEG) linkage or four consecutive deoxythymidines together with an inverted deoxythymidine nucleotide at the 3′-end. The anticoagulation potential, the reversal of coagulation with different antidote sequences, and the nuclease stability of the aptamers were then investigated. The results showed that a bivalent aptamer RNV220 containing an inverted deoxythymidine and a TEG linkage chemistry significantly enhanced the anticoagulation properties in blood plasma and nuclease stability compared to the existing aptamer designs. Furthermore, a bivalent antidote sequence RNV220AD efficiently reversed the anticoagulation effect of RNV220 in blood plasma. Based on our results, we believe that RNV220 could be developed as a potential anticoagulant therapeutic molecule. MDPI 2017-10-19 /pmc/articles/PMC6151750/ /pubmed/29048375 http://dx.doi.org/10.3390/molecules22101770 Text en © 2017 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (http://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Hughes, Quintin W.
Le, Bao T.
Gilmore, Grace
Baker, Ross I.
Veedu, Rakesh N.
Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties
title Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties
title_full Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties
title_fullStr Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties
title_full_unstemmed Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties
title_short Construction of a Bivalent Thrombin Binding Aptamer and Its Antidote with Improved Properties
title_sort construction of a bivalent thrombin binding aptamer and its antidote with improved properties
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC6151750/
https://www.ncbi.nlm.nih.gov/pubmed/29048375
http://dx.doi.org/10.3390/molecules22101770
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